LOCUS       sequence-335632-        7005 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  FKBP anchor for retaining DHFR-fused cargoes in the ER in the 
            presence of zapalog. Encodes ER retrieval motif (KDEL) at C-terminus
            of mCherry. Contains IRES following ORF for cloning cargo. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7005)
  AUTHORS   Bourke AM, Schwartz SL, Bowen AB, Kleinjan MS, Winborn CS, Kareemo 
            DJ, Gutnick A, Schwarz TL, Kennedy MJ
  TITLE     zapERtrap: A light-regulated ER release system reveals unexpected 
            neuronal trafficking pathways.
  JOURNAL   J Cell Biol. 2021 Sep 6;220(9). pii: 212461. doi: 
            10.1083/jcb.202103186. Epub 2021 Jul 9.
  PUBMED    34241635
REFERENCE   2  (bases 1 to 7005)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Cell 
            Biol."; date: "2021-09-6"; volume: "220(9). pii: 212461. doi"; 
            pages: " 10.1083/jcb.202103186. Epub 2021 Jul 9"
COMMENT     Sequence Label: 2xFKBP-mCh-KDEL
FEATURES             Location/Qualifiers
     source          1..7005
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(42..630)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(122..141)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(801..1592,1593..1661))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     1424..1443
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1662..1766)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        1797..2176
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2178..2455
                     /label=chicken beta-actin promoter
     intron          2456..3468
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     3392..3414
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     3476..3495
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     sig_peptide     3529..3588
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     CDS             3592..3618
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3625..3945
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     CDS             3973..4293
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     regulatory      4330..4339
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4336..5043
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(4481..4499)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     4749..4768
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    5083..5656
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(5250..5267)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     5477..5496
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     complement(5789..5808)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    5854..5909
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(5908..5927)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(6300..6316)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6300..6316)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6313..6335)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6324..6340
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(6348..6354,6355..6372,6373..6378))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6393..6414
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        6473..6668
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      6519..6654
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    6674..6808
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6711..6730)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6765..6784
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6876..6893)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
       61 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
      121 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
      181 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
      241 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
      301 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
      361 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
      421 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
      481 agracagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
      541 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
      601 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
      661 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
      721 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
      781 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
      841 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
      901 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
      961 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     1021 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     1081 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     1141 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     1201 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     1261 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     1321 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     1381 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     1441 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     1501 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     1561 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     1621 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     1681 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     1741 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgggtcgaca
     1801 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
     1861 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
     1921 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
     1981 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
     2041 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
     2101 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
     2161 catcgctatt accatggtcg aggtgagccc cacgttctgc ttcactctcc ccatctcccc
     2221 cccctcccca cccccaattt tgtatttatt tattttttaa ttattttgtg cagcgatggg
     2281 ggcggggggg gggggggggc gcgcgccagg cggggcgggg cggggcgagg ggcggggcgg
     2341 ggcgaggcgg agaggtgcgg cggcagccaa tcagagcggc gcgctccgaa agtttccttt
     2401 tatggcgagg cggcggcggc ggcggcccta taaaaagcga agcgcgcggc gggcgggagt
     2461 cgctgcgttg ccttcgcccc gtgccccgct ccgcgccgcc tcgcgccgcc cgccccggct
     2521 ctgactgacc gcgttactcc cacaggtgag cgggcgggac ggcccttctc ctccgggctg
     2581 taattagcgc ttggtttaat gacggctcgt ttcttttctg tggctgcgtg aaagccttaa
     2641 agggctccgg gagggccctt tgtgcggggg ggagcggctc ggggggtgcg tgcgtgtgtg
     2701 tgtgcgtggg gagcgccgcg tgcggcccgc gctgcccggc ggctgtgagc gctgcgggcg
     2761 cggcgcgggg ctttgtgcgc tccgcgtgtg cgcgagggga gcgcggccgg gggcggtgcc
     2821 ccgcggtgcg ggggggctgc gaggggaaca aaggctgcgt gcggggtgtg tgcgtggggg
     2881 ggtgagcagg gggtgtgggc gcggcggtcg ggctgtaacc cccccctgca cccccctccc
     2941 cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtacggggcg tggcgcgggg
     3001 ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc cgggcggggc ggggccgcct
     3061 cgggccgggg agggctcggg ggaggggcgc ggcggccccc ggagcgccgg cggctgtcga
     3121 ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc gcagggactt
     3181 cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac cccctctagc
     3241 gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga gggccttcgt
     3301 gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc gcggggggac
     3361 ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt gaccggcggc
     3421 tctagagcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct cctgggcaac
     3481 gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat tgcccaccat ggagacagac
     3541 acactcctgc tatgggtact gctgctctgg gttccaggtt ccactggtga ctatccatat
     3601 gatgttccag attatgctct cgagggagtc caggtagaga cgatttcacc cggcgacggc
     3661 cgcacattcc cgaagcgagg ccagacatgt gtggtgcatt ataccgggat gcttgaggat
     3721 ggaaagaagt tcgactcaag tagggatcgg aacaaaccgt tcaagttcat gttgggaaag
     3781 caagaagtaa tacggggttg ggaggaaggt gttgcccaaa tgagtgttgg tcagagagct
     3841 aagctgacga tcagtcccga ctatgcctat ggtgcaacag gacatccagg aataatacct
     3901 ccgcacgcca ctttggtatt cgatgtcgaa ctcttgaagc tggaaagtgc tggtagtgct
     3961 ggtagtgctg gtggagtgca ggtggaaacc atctccccag gagacgggcg caccttcccc
     4021 aagcgcggcc agacctgcgt ggtgcactac accgggatgc ttgaagatgg aaagaaattt
     4081 gattcctccc gggacagaaa caagcccttt aagtttatgc taggcaagca ggaggtgatc
     4141 cgaggctggg aagaaggggt tgcccagatg agtgtgggtc agagagccaa actgactata
     4201 tctccagatt atgcctatgg tgccactggg cacccaggca tcatcccacc acatgccact
     4261 ctcgtcttcg atgtggagct tctaaaactg gaaggtggcg gcggaagcgg tggcgggggt
     4321 ggaccggtcg ccaccatggt gagcaagggc gaggaggata acatggccat catcaaggag
     4381 ttcatgcgct tcaaggtgca catggagggc tccgtgaacg gccacgagtt cgagatcgag
     4441 ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag
     4501 ggtggccccc tgcccttcgc ctgggacatc ctgtcccctc agttcatgta cggctccaag
     4561 gcctacgtga agcaccccgc cgacatcccc gactacttga agctgtcctt ccccgagggc
     4621 ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg tggtgaccgt gacccaggac
     4681 tcctccctgc aggacggcga gttcatctac aaggtgaagc tgcgcggcac caacttcccc
     4741 tccgacggcc ccgtaatgca gaagaagacc atgggctggg aggcctcctc cgagcggatg
     4801 taccccgagg acggcgccct gaagggcgag atcaagcaga ggctgaagct gaaggacggc
     4861 ggccactacg acgctgaggt caagaccacc tacaaggcca agaagcccgt gcagctgccc
     4921 ggcgcctaca acgtcaacat caagttggac atcacctccc acaacgagga ctacaccatc
     4981 gtggaacagt acgaacgcgc cgagggccgc cactccaccg gcggcatgga cgagctgtac
     5041 aagaaggatg agctgtaaac ggtaccgcgg gcccgggatc cgcccctctc cctccccccc
     5101 ccctaacgtt actggccgaa gccgcttgga ataaggccgg tgtgcgtttg tctatatgtt
     5161 attttccacc atattgccgt cttttggcaa tgtgagggcc cggaaacctg gccctgtctt
     5221 cttgacgagc attcctaggg gtctttcccc tctcgccaaa ggaatgcaag gtctgttgaa
     5281 tgtcgtgaag gaagcagttc ctctggaagc ttcttgaaga caaacaacgt ctgtagcgac
     5341 cctttgcagg cagcggaacc ccccacctgg cgacaggtgc ctctgcggcc aaaagccacg
     5401 tgtataagat acacctgcaa aggcggcaca accccagtgc cacgttgtga gttggatagt
     5461 tgtggaaaga gtcaaatggc tctcctcaag cgtattcaac aaggggctga aggatgccca
     5521 gaaggtaccc cattgtatgg gatctgatct ggggcctcgg tacacatgct ttacatgtgt
     5581 ttagtcgagg ttaaaaaaac gtctaggccc cccgaaccac ggggacgtgg ttttcctttg
     5641 aaaaacacga tgataatgaa ttctgcagtc gacggtaccg cgggccctac aagtaaagcg
     5701 gccgccaatt cactcctcag gtgcaggctg cctatcagaa ggtggtggct ggtgtggcca
     5761 atgccctggc tcacaaatac cactgagatc tttttccctc tgccaaaaat tatggggaca
     5821 tcatgaagcc ccttgagcat ctgacttctg gctaataaag gaaatttatt ttcattgcaa
     5881 tagtgtgttg gaattttttg tgtctatcac tcggaaggac atatgggagg gcaaatcatt
     5941 taaaacatca gaatgagtat ttggtttaga gtttggcaac atatgcccat atgctggctg
     6001 ccatgaacaa aggttggcta taaagaggtc atcagtatat gaaacagccc cctgctgtcc
     6061 attccttatt ccatagaaaa gccttgactt gaggttagat tttttttata ttttgttttg
     6121 tgttattttt ttctttaaca tccctaaaat tttccttaca tgttttacta gccagatttt
     6181 tcctcctctc ctgactactc ccagtcatag ctgtccctct tctcttatgg agatccctcg
     6241 acctgcaccg tcgaccagct ggtcgacggt gcaccgtcga ccagcttggc gtaatcatgg
     6301 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     6361 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     6421 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagcggat ccgcatctca
     6481 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     6541 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     6601 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     6661 tttgcaaaaa gctaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     6721 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     6781 catcaatgta tcttatcatg tctggatccg ctgcattaat gaatcggcca acgcgcgggg
     6841 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     6901 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     6961 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccag
//
